Ras proteins are key molecular switches that couple extracellular stimuli to intracellular response machinery. Oncogenic mutations in the N- and K-ras genes are frequently identified in patients with myeloid eukemia. Despite intensive investigation the roles of oncogenic Ras mutations in cell proliferation, lineage differentiation, and leukemogenesis are still not fully understood, nor are the outputs of individual signaling pathways fully known. Previous studies showed that overexpression of oncogenic Ras leads to abnormal proliferation and blockage of terminal differentiation of erythroid progenitors, and that these effects are mediated mainly through constitutive activation of the MEK/MAPK pathway. However, when expressed at a physiological level, oncogenic K-ras does not constitutively activate but rather hyperactivates two downstream signaling pathways in response to cytokine stimulation: the MEK/MAPK and PI3K/Akt pathways. This fellowship will probe the cellular and molecular mechanisms that underlie how oncogenic K-ras signaling, at its physiological level, predisposes hematopoietic cells to leukemia. To accomplish this, biochemical, cellular, molecular, and mouse genetic approaches will be used. In the proposed experimental system, expression of oncogenic K-ras from its endogenous locus is induced in primary hematopoietic progenitors. Studies of the amplitude and duration of cytokine-dependent signaling in these cells will unravel how expression of oncogenic K-ras changes the cellular responses to different cytokine stimulations. Moreover, examination of the outputs of individual signaling pathways downstream of oncogenic K-ras will reveal their individual roles in regulating cell proliferation and differentiation. Together, these results will establish the sufficiency and necessity of individual signaling pathways in oncogenic K-ras-mediated effects. Importantly, analysis of the transcriptional profiles in wild-type hematopoietic progenitors as well as in progenitors expressing abnormal signaling proteins will provide gene-expression signatures, that are indicative of potential targets of oncogenic K-ras signaling as well as potential targets of individual signaling pathways in human myeloid leukemia. This analysis will shed light on the mechanisms underlying leukemogenesis and leukemia progression. Deregulation of Ras signaling pathways resulting from oncogenic mutations in ras genes or their regulators have been found in all human cancer types. Therefore, studying the cellular and molecular mechanisms by which oncogenic K-ras controls cell proliferation and lineage differentiation might identify new drug targets or therapeutic approaches to treat leukemia as well as other cancer types. Through this proposed research, the applicant will receive training in bioinformatics, functional genomics and mouse genetics and will gain the necessary skills and knowledge to become an independent investigator.